1. Field of the Invention
The invention relates to compounds that are capable of inhibiting sphingosine kinase and that are useful for treating or preventing hyperproliferative disease, autoimmune disease, inflammatory disease, or allergy. The invention also relates to methods and compositions for treating or preventing hyperproliferative disease, autoimmune disease, inflammatory disease, or allergy.
2. Description of the Related Art
The mechanisms and effects of the interconversion of sphingolipids have been the subjects of a growing body of scientific investigation. Sphingomyelin, for example, is not only a building block for cellular membranes but also serves as the precursor for potent lipid messengers that have profound cellular effects. Stimulus-induced metabolism of these lipids is critically involved in cancer cell biology. Consequently, these metabolic pathways offer exciting new molecular targets for the development of anticancer drugs.
As depicted in FIG. 1, ceramide is produced by the hydrolysis of sphingomyelin in response to stresses including chemotherapeutic drugs. Ceramide induces apoptosis in proliferating cells by a mechanism that remains to be elucidated. However, ceramide can be further hydrolyzed by the action of ceramidase to produce sphingosine, which is then rapidly phosphorylated by sphingosine kinase (SK) in many different cell types to produce sphingosine-1-phosphate (S1P). The actions of ceramidase and SK appear to be rapidly activated by a number of growth factors. A critical balance between the concentrations of ceramide and S1P, termed the ceramide/S1P rheostat, has been hypothesized to determine the fate (proliferation or apoptosis) of the cell. In this view, it is the balance between the cellular concentrations of ceramide and S1P that determines whether a cell proliferates or undergoes apoptosis.
Upon exposure to mitogens or intracellular oncoproteins, cells experience a rapid increase in the intracellular levels of S1P and depletion of ceramide levels, and consistent with the hypothesis, this “setting” of the rheostat promotes cell survival and proliferation. In contrast, activation of sphingomyelinase in the absence of activation of ceramidase and SK results in the accumulation of ceramide and subsequent apoptosis. Importantly, ceramide appears to induce apoptosis in tumor cells without disrupting quiescent normal cells. Furthermore, ceramide enhances apoptosis in response to anticancer drugs including Taxol and etoposide.
Accumulating evidence confirms that S1P is a critical second messenger that exerts proliferative and antiapoptotic actions. For example, microinjection of S1P into mouse oocytes induces DNA synthesis, and promotes the secretion of Insulin-like Growth Factor-II (IGF-II) from human breast carcinoma cells. Additionally, it has been shown that S1P effectively inhibits ceramide-induced apoptosis in association with decreased caspase activation. These studies in various cell lines consistently indicate that S1P is able to induce proliferation and protect cells from ceramide-induced apoptosis. While the elucidation of downstream targets of S1P remains an interesting problem in cell biology, sufficient validation of these pathways has been established to justify their evaluation as targets for new types of anticancer drugs. As S1P appears to be the most direct mitogenic messenger, inhibition of its production should have profound antiproliferative effects on tumor cells.
Sphingosine kinase (SK) is the sole enzyme responsible for S1P production in cells. The enzyme was initially isolated from rat kidney, and demonstrated KM values 5 μM and 93 μM for sphingosine and ATP, respectively. The human isoform was cloned in 2000, and displays similar physical and biochemical characteristics. Shortly thereafter, a second SK isoform was cloned: however, this species displays much lower activity and different kinetic profiles than the type 1 enzyme. RNA encoding SK is detected in most tissues, with higher levels in lung and spleen. Interestingly, a number of studies have shown that a variety of proliferative factors, including protein kinase C (PKC) activators, fetal calf serum and platelet-derived growth factor, epidermal growth factor (EGF), and tumor necrosis factor-alpha (TNF-α) rapidly elevate cellular SK activity.
Recently, an oncogenic role of SK has been directly demonstrated. In these studies, transfection of SK into NIH 3T3 fibroblasts was sufficient to promote foci formation and cell growth in soft-agar, and to allow these cells to form tumors in NOD/SCID mice. Additionally, inhibition of SK by transfection with a dominant-negative SK mutant or by treatment of cells with the nonspecific SK inhibitor D-erythro-N,N-dimethylsphingosine blocked transformation mediated by oncogenic H-Ras. Since abnormal activation of Ras as well as overexpression and mutation of ras family genes frequently occurs in cancer, these findings suggest a significant role of SK in this disease. Another study showed that a cellular receptor that specifically binds S1P, termed EDG4, is a specific marker for ovarian cancer cells. S1P has also been implicated in angiogenesis, as it induces motility and mitogenesis in smooth muscle cells and endothelial cell differentiation. These various findings indicate that SK is an important molecular target in cancer.
Despite the high level of interest in sphingolipid-derived signaling, there are very few demonstrated inhibitors of the enzymes of this pathway. In particular, the field suffers from a lack of potent and selective inhibitors of SK. Pharmacological studies to date have used three compounds to inhibit SK activity: D-erythro-N,N-dimethylsphingosine, D,L-threo-dihydrosphingosine and N,N,N-trimethyl-sphingosine. However, these compounds are not specific inhibitors of SK and have been shown to affect other important cellular proteins, including PKC, sphingosine-dependent protein kinase, 3-phosphoinositide-dependent kinase, and casein kinase II. Therefore, there is a need in the art for selective and potent inhibitors of SK as antiproliferative and anti-inflammatory agents.